Abstract
INTRODUCTION: Alpha-linolenic acid (ALA), an essential polyunsaturated fatty acid, modulates gastrointestinal microbiota and host immunity, yet its regulatory mechanisms in dairy cows remain unclear. METHODS: This study investigated how dietary ALA influenced gut microbiota, metabolome, and immune function in lactating Holstein cows. Ten cows were randomly assigned to two groups (n = 5) receiving either a low-ALA (LALA, 5.02 ± 0.09% ALA of total fatty acids) or high-ALA diet (HALA, 32.04 ± 1.55% ALA of total fatty acids). Rumen fluid, feces, and blood samples were analyzed post-intervention. RESULTS: The HALA group exhibited increased ruminal abundance of Eubacterium coprostanoligenes group and Ruminococcus (p < 0.05), alongside reduced proinflammatory metabolites including dodecanoic acid, myristic acid, and prostaglandin I(2) in the rumen. Plasma leukotriene C(4) levels were also decreased (p < 0.05). Metabolomic enrichment analysis revealed significant downregulation of arachidonic acid metabolism. Correlation analyses demonstrated that Eubacterium coprostanoligenes group negatively associated with suppressed prostaglandin I(2) (rumen metabolite) and leukotriene C(4) (plasma metabolite), but positively correlated with enriched fecal Clostridia UCG-014 and Ruminococcus. DISCUSSION: These findings indicate that high dietary ALA reshapes gastrointestinal microbiota and attenuates inflammatory responses by inhibiting microbial-metabolite-driven arachidonic acid metabolism, thereby enhancing immune regulation in dairy cows.